LOCUS       sequence-477568-        5732 bp    DNA     circular SYN 02-JUL-2025
DEFINITION  Tet inducible PFE3 for ablating excitatory synapses..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5732)
  AUTHORS   Bareghamyan A, Deng C, Daoudi S, Yadav SC, Lu X, Zhang W, Campbell 
            RE, Kramer RH, Chenoweth DM, Arnold DB
  TITLE     A toolbox for ablating excitatory and inhibitory synapses.
  JOURNAL   eLife. 2025 Apr 29;13:RP103757. doi: 10.7554/eLife.103757.
  PUBMED    40298577
REFERENCE   2  (bases 1 to 5732)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 2, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "eLife."; 
            date: "2025-04-29"; volume: "13:RP103757. doi"; pages: " 
            10.7554/eLife.103757"
COMMENT     Sequence Label: TRE-PFE3-HA
FEATURES             Location/Qualifiers
     source          1..5732
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        31..409
                     /label=TRE3G promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G"
     protein_bind    39..57
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    75..93
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    111..129
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    147..165
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    183..201
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    219..237
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    255..273
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     318..342
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             696..722
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     intron          1624..1756
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1739..1758
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    1765..2353
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1818..1838)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2236..2247)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2380..2397)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2386..2497
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2593..2613)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2593..2611)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2632..2648)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2754..2773)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2819..2874
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2873..2892)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3235..3251)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3259..3275
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(3283..3289,3290..3307,3308..3313))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3328..3349
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3408..3603
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3454..3589
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3609..3743
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3646..3665)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3700..3719
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3811..3828)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3982..4570)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4062..4081)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4741..5532,5533..5601))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5364..5383
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5602..5706)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgacatt gattattgac tagttatctc gagtttactc cctatcagtg atagagaacg
       61 tatgaagagt ttactcccta tcagtgatag agaacgtatg cagactttac tccctatcag
      121 tgatagagaa cgtataagga gtttactccc tatcagtgat agagaacgta tgaccagttt
      181 actccctatc agtgatagag aacgtatcta cagtttactc cctatcagtg atagagaacg
      241 tatatccagt ttactcccta tcagtgatag agaacgtata agctttaggc gtgtacggtg
      301 ggcgcctata aaagcagagc tcgtttagtg aaccgtcaga tcgcctggag caattccaca
      361 acacttttgt cttataccaa ctttccgtac cacttcctac cctcgtaaag tcgacaccgg
      421 ggcccatgct cgaagtcaag gaagcatcac caaccagcat ccagatcagc tgggtgctcc
      481 acttgcgcca cgttcgctac taccgcatca cctacggtga aactggtggc aatagccctg
      541 tccaggaatt caccgtgcct ggcagcaagt ccactgctac catcagcggc ctgaaacctg
      601 gtgtcgacta taccatcacg gtgtacgccg tcacgatctt cagcgcctac cgctccgcct
      661 ggccgccgat ctccatcaac taccgcacca ccggttaccc atacgatgtt ccagattacg
      721 ctggtggaag cacgcgttgt gtgatttgcc aggggcggcc taaaaatggt tgcattgttc
      781 acggcaaaac cggacacctg atgtcatgtt tcacgtgtgc aaaggcacta gctgcggcga
      841 atgcgccctg ccccgtgtgc agagctagca agctctcagg tggcggaggg agcggaggag
      901 gtggctctgg aggtggtggc tcagctgaag catggtacaa tctgggaaac gcttattaca
      961 agcagggcga ctatcaaaaa gccattgagt attaccaaaa ggctcttgag ctagacccta
     1021 ataacgccga agcgtggtat aacctaggca acgcatatta caagcaggga gactaccaaa
     1081 aggcaatcga atattaccag aaggcactcg aactggaccc aaataacgca gaggcttggt
     1141 ataatctggg aaacgcctac tataaacagg gtgactacca aaaggcgata gaggactatc
     1201 agaaggcctt ggagctcgat ccgaacaatc tgcaacgtag tgctggtggc ggagggtctg
     1261 gtggaggagg ctcaggagga ggtggcgcat caggtaccaa gaagaaactc agcaagtcag
     1321 acatcatgtt ggtgcagagc gccaatgtcc ctagcaaccc gacccaggtg ccggtagagg
     1381 agtccggagg cttcggttcc catcaccaca atcagaacta ttgctatcag gtctgcctca
     1441 ccccagagtc cgccaagacc gacctgatgt tcttgaagcc ctgcagccct tcccggagta
     1501 cagacgctga gcacaatccc tgtggagcca tcgtcaccgg gtacagcgac cagcaaccag
     1561 acatcatttc caacggaagc attttgtcca acgagaatta aatcgattat gtcgaggccg
     1621 caggtaagta tcaaggttac aagacaggtt taaggagacc aatagaaact gggcttgtcg
     1681 agacagagaa gactcttgcg tttctgatag gcacctattg gtcttactga catccacttt
     1741 gcctttctct ccacaggtgt cgacaatcaa cctctggatt acaaaatttg tgaaagattg
     1801 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     1861 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     1921 ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact
     1981 gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc
     2041 gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc
     2101 cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaag
     2161 ctgacgtcct ttccatggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc
     2221 ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg
     2281 gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg
     2341 gccgcctccc cgcctggaat tcgagctcgg tacgatcagc ctcgactgtg ccttctagtt
     2401 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     2461 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     2521 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagcc
     2581 cagcttttgt tccctttagt gagggttaat tgcgcgcttg gcgtaatcat ggtcatagct
     2641 gtttcctgtg tgaaattgtt atccgctaat tcactcctca ggtgcaggct gcctatcaga
     2701 aggtggtggc tggtgtggcc aatgccctgg ctcacaaata ccactgagat cgatcttttt
     2761 ccctctgcca aaaattatgg ggacatcatg aagccccttg agcatctgac ttctggctaa
     2821 taaaggaaat ttattttcat tgcaatagtg tgttggaatt ttttgtgtct ctcactcgga
     2881 aggacatatg ggagggcaaa tcatttaaaa catcagaatg agtatttggt ttagagtttg
     2941 gcaacatatg cccatatgct ggctgccatg aacaaaggtt ggctataaag aggtcatcag
     3001 tatatgaaac agccccctgc tgtccattcc ttattccata gaaaagcctt gacttgaggt
     3061 tagatttttt ttatattttg ttttgtgtta tttttttctt taacatccct aaaattttcc
     3121 ttacatgttt tactagccag atttttcctc ctctcctgac tactcccagt catagctgtc
     3181 cctcttctct tatggagatc cctcgacctg cagcccaagc ttggcgtaat catggtcata
     3241 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     3301 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     3361 ctcactgccc gctttccagt cgggaaacct gtcgtgccag cggatccgca tctcaattag
     3421 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     3481 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     3541 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     3601 aaaaagctaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     3661 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     3721 atgtatctta tcatgtctgg atccgctgca ttaatgaatc ggccaacgcg cggggagagg
     3781 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     3841 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     3901 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     3961 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     4021 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     4081 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     4141 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     4201 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     4261 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     4321 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     4381 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     4441 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     4501 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     4561 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     4621 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     4681 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     4741 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     4801 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     4861 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     4921 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     4981 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     5041 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     5101 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     5161 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     5221 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     5281 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     5341 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     5401 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     5461 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     5521 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     5581 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     5641 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     5701 tccgcgcaca tttccccgaa aagtgccacc tg
//
